Mucosal Cells Research Articles

LRRC8A drives NADPH oxidase-mediated mitochondrial dysfunction and inflammation in allergic rhinitis.

Allergic rhinitis (AR) is a complex disorder with variable pathogenesis. Increasing evidence suggests that the LRRC8A is involved in maintaining cellular stability, regulating immune cell activation and function, and playing significant roles in inflammation. However, the involvement of LRRC8A in AR inflammation and its underlying mechanisms remain unclear. LRRC8A expression in AR patients, confirmed by qRT-PCR and Western blotting, was analyzed to investigate its relationship with the clinical characteristics of AR patients. In vitro, IL-13 stimulated HNEpCs to establish a Th2 inflammation model, with subsequent LRRC8A knockout or overexpression. NOX1/NOX4 inhibitor (GKT137831) and chloride channel inhibitor (DCPIB) were utilized to investigate AR development mechanisms during LRRC8A overexpression. An OVA-induced AR model with nasal mucosa LRRC8A knockdown confirmed LRRC8A's regulatory role in AR inflammation. LRRC8A mRNA and protein levels were significantly elevated in AR patients, positively correlating with NADPH oxidase subunits and Th2 inflammatory markers. In vitro, IL-13 stimulation of HNEpCs resulted in upregulation of LRRC8A and increased expression of NOX1, NOX4, and p22phox, along with mitochondrial dysfunction and NF-κB pathway activation. The knockout of LRRC8A reversed these effects. In nasal mucosal epithelial cells, DCPIB and GKT137831 completely blocked mitochondrial dysfunction caused by the overexpression of LRRC8A, which led to up-regulation of NOX1, NOX4, and p22phox. In vivo, knocking down LRRC8A reduced eosinophil infiltration, downregulated the expression of NOX1, NOX4, p22phox IL-4, IL-5, and IL-13, and decreased NF-κB pathway activation. LRRC8A drives the upregulation of NOX1, NOX4, and p22phox, leading to ROS overproduction and mitochondrial dysfunction. It also activates NF-κB, ultimately leading to nasal mucosal epithelial inflammation. LRRC8A may be a potential target for the treatment of AR.

Mechanosensitive release of ATP in the urinary bladder mucosa.

The urinary bladder mucosa (urothelium and suburothelium/lamina propria) functions as a barrier between the content of the urine and the underlying bladder tissue. The bladder mucosa is also a mechanosensitive tissue that releases signaling molecules that affect functions of cells in the bladder wall interconnecting the mucosa with the detrusor muscle and the CNS. Adenosine 5'-triphosphate (ATP) is a primary mechanotransduction signal that is released from cells in the bladder mucosa in response to bladder wall distention and activates cell membrane-localized P2X and P2Y purine receptors on urothelial cells, sensory and efferent neurons, interstitial cells, and detrusor smooth muscle cells. The amounts of ATP at active receptor sites depend significantly on the amounts of extracellularly released ATP. Spontaneous and distention-induced release of ATP appear to be under differential control. This review is focused on mechanisms underlying urothelial release of ATP in response to mechanical stimulation. First, we present a brief overview of studies that report mechanosensitive ATP release in bladder cells or tissues. Then, we discuss experimental evidence for mechanosensitive release of urothelial ATP by vesicular and non-vesicular mechanisms and roles of the stretch-activated channels PIEZO channels, transient receptor potential vanilloid type 4, and pannexin 1. This is followed by brief discussion of possible involvement of calcium homeostasis modulator 1, acid-sensing channels, and connexins in the release of urothelial ATP. We conclude with brief discussion of limitations of current research and of needs for further studies to increase our understanding of mechanotransduction in the bladder wall and of purinergic regulation of bladder function.

Association between sleep quality and polymorphisms of the genes COMT, HTR2A and FKBP5 in individuals with and without dentofacial deformity

Sleep is a fundamental biological function, and any disturbances can lead to alterations in an individual’s physical, occupational, cognitive, and social functioning. Aim: This study aimed to evaluate the quality of sleep and its association with factors such as age, sex, facial profile, and genetic polymorphisms in individuals who underwent dental treatment. Methods: The study comprised a total of 227 individuals. The “Sleep Assessment Questionnaire” was utilized to evaluate sleep quality. For the genetic assessment, oral mucosa cells were collected and markers from the COMT (rs174675, rs165656), HTR2A (rs4941573, rs6313), and FKBP5 (rs1360780, rs3800373) genes were selected for real-time PCR analysis. The data were subjected to statistical analysis with a significance level of 0.05. Results: The results showed that women had a poorer perception of sleep quality (p<0.05). There was a significant association between sleep quality and facial profile (p<0.05). Individuals with facial profiles I and II had a poorer perception of sleep quality in general and in the domains of non-restorative sleep, sleep time disturbance, and restlessness. Individuals with facial profile II had a poorer perception of sleep apnea than those with profile III (p=0.034). There was a significant association between the COMT rs174675 polymorphism and restlessness (p=0.035). Conclusion: The poorest perception of sleep quality was associated with women and individuals with facial profiles I and II. The poorest perception of restlessness was associated with COMT rs174675 polymorphism.

Comprehensive gene expression analysis in gallbladder mucosal epithelial cells of dogs with gallbladder mucocele.

Gallbladder mucocele (GBM) is a common disease in the canine gallbladder. Although the pathogenesis of GBM remains unclear, we recently reported that the excessive accumulation of mucin in the gallbladder is not a result of overproduction by gallbladder epithelial cells (GBECs). Changes in the function of GBECs other than the production of mucin are associated with the pathogenesis of GBM. We performed an RNA sequencing (RNA-seq) analysis to comprehensively search for abnormalities in gene expression profiles of GBECs in dogs with GBM. Fifteen dogs with GBM and 8 dogs euthanized for reasons other than gallbladder disease were included. The GBECs were isolated from gallbladder tissues to extract RNA. The RNA-seq analysis was performed using the samples from 3 GBM cases and 3 dogs with normal gallbladders, and the gene expression profiles were compared between the 2 groups. Differences in mRNA expression levels of the extracted differentially expressed genes (DEGs) were validated by quantitative reverse transcription polymerase chain reaction (RT-qPCR) using samples of 15 GBM cases and 8 dogs with normal gallbladders. Comparison of gene expression profiles by RNA-seq extracted 367 DEGs, including ANO1, a chloride channel associated with changes in mucin morphology, and HTR4, which regulates the function of chloride channels. The ANO1 and HTR4 genes were confirmed to be downregulated in the GBM group by RT-qPCR. Our results suggest that GBM may be associated with decreased function of chloride channels expressed in GBECs.

The Impact of NO2 on Epithelial Barrier Integrity of a Primary Cell-Based Air-Liquid Interface Model of the Nasal Respiratory Epithelium.

Nitrogen dioxide (NO2) is a pervasive gaseous air pollutant with well-documented hazardous effects on health, necessitating precise toxicological characterization. While prior research has primarily focused on lower airway structures, the upper airways, serving as the first line of defense against airborne substances, remain understudied. This study aimed to investigate the functional effects of NO2 exposure alone or in combination with hypoxia as a secondary stimulus on nasal epithelium and elucidate its molecular mechanisms because hypoxia is considered a pathophysiological factor in the onset and persistence of chronic rhinosinusitis, a disease of the upper airways. Air-liquid interface cell cultures derived from primary nasal mucosa cells were utilized as an invitro model, offering a high invitro-in vivo correlation. Our findings demonstrate that NO2 exposure induces malfunction of the epithelial barrier, as evidenced by decreased transepithelial electrical resistance and increased fluorescein isothiocyanate (FITC)-dextran permeability. mRNA expression analysis revealed a significant increase in IL-6 and IL-8 expressions following NO2. Reduced mRNA expression of the tight junction component occludin was identified as a structural correlate of the damaged epithelial barrier. Notably, hypoxic conditions alone did not alter epithelial barrier integrity. These findings provide information on the harmful effects of NO2 exposure on the human nasal epithelium, including compromised barrier integrity and induction of inflammatory responses. Overall, this study contributes to our understanding of pathophysiological mechanisms underlying also upper airway respiratory diseases associated with air pollution exposure and emphasizes the importance of mitigating NO2 emissions to safeguard respiratory health.

Immune cell changes in Helicobacter pylori infection-induced glandular epithelial cell damage of the gastric mucosa

Objective The purpose of this study was to investigate the relationship between Helicobacter pylori (H. pylori) infection-induced changes in gastric mucosal immune cells and glandular epithelial cell damage and the histopathological characteristics of these changes. Methods We performed a detailed histomorphometry and immunohistochemical analysis of a total of 1635 H. pylori-infected gastric mucosal specimens. Results Stage-wise features were as follows: Early stage of infection: H. pylori was colonized in the mucous layer, and very few neutrophils were visible in the layer. Gastric surface epithelial cell infection stage: H. pylori specifically and selectively adhered to the cytoplasm of the surface mucous cells, with the presence of a small number of neutrophils, eosinophils, and lymphocytes infiltration, which is termed early immune response. Compensatory mucous neck cell hyperplasia stage: proliferation of stem cells in the deep gastric pit and infiltration of a large number of lymphocytes in the superficial layer of lamina propria were observed, which is termed immune cell mobilization counterinsurgency. Lamina propria lesion stage: excessive upward migration of the proliferative area. Infiltration of a large number of lymphocytes, plasma cells, monocytes, macrophages, and other immune cells was observed in the whole layer of the gastric mucosa, which is termed automatic replication of immune cells. Abnormal proliferative transformation stage: presence of intranuclear milky globular body cells or signet-ring cell-like heterotypic cells at the junction of the gastric pit and the gastric gland, with proliferation of large numbers of immune cells and vacuolar degeneration of immune cells, which is termed the proliferation and degeneration of immune cells. Intraepithelial neoplasia stage: clonal hyperplasia of epithelial cells and immunosuppression. Conclusion For controlling the occurrence and development of gastric cancer and effective immune intervention, it is essential to grasp the relationship between H. pylori infection-induced changes in gastric mucosal immune cells and glandular epithelial cell damage and its histopathological characteristics.

Prevalence of human papilloma virus in head and neck mucous squamous cell carcinoma and genotypes by location: an observational study.

Human papillomavirus (HPV) is a factor in oropharyngeal cancer, but data regarding other head and neck locations are scarce in France. The main objective of the study was to determine the prevalence of HPV in head and neck cancers at all locations. As a secondary objective, we aimed to investigate the HPV genotypes. We retrospectively included in a tertiary center between 2014 and 2020 mucosal squamous cell carcinomas of the head and neck in adult. First outcome was the prevalence of HPV cancer. Secondary outcomes were overall survival (OS) at 2 and 5 years and disease-free survival (DFS). A total of 508 patients were enrolled, resulting in 537 cases of mucous squamous cell carcinoma of the head and neck (n = 29 synchronous carcinomas). Clinical, pathological, and survival data were collected, and a double PCR for HPV with genotyping was performed on most of the samples. The HPV prevalence in the cohort was 28.2%, with HPV 16 being the predominant genotype (87%). However, HPV-positive status did not significantly improve OS at 2 and 5 years or DFS (P = 0.1, P = 0.64, and P = 0.07, respectively). It was also observed that HPV-positive patients had significantly fewer second tumor localizations (P < 0.01). The prevalence of HPV continues to rise, and the complexities surrounding HPV status and its association with clinical outcomes in head and neck squamous cell carcinoma highlight the impact of vaccination.

Dual-Modality Accurate Visualization of Drug Synergy Based on Mass Spectrometry and Fluorescence Imaging.

There is a potential synergistic effect between nonsteroidal anti-inflammatory drugs and hydrogen sulfide (H2S), but direct evidence for the study is lacking. With a single fluorescence detection method, it is difficult to accurately confirm the effectiveness of the synergistic effect. In this study, the fluorescent probe and the nonsteroidal anti-inflammatory drug naproxen were combined via different self-immolative spacer groups to obtain a diagnostic and therapeutic integrated fluorescent probe Nap-NP-NSB, which can release H2S. The quantitative release of H2S by Nap-NP-NSB was evaluated in vitro and in cells, and the synergistic effect of H2S and naproxen was confirmed by monitoring the treatment process of cellular inflammation and oxidative damage of gastric mucosa cells. Finally, in vivo fluorescence imaging and mass spectrometry imaging of the liver and stomach tissues and their sections were performed in the mouse model of acute hepatitis. The dual-modal detection method not only confirmed that Nap-NP-NSB had better anti-inflammatory activity and less gastric mucosal damage, but also enabled a more accurate visualization of the drug synergistic effect of naproxen and H2S. This work provides a dual visualization imaging method combining fluorescence and mass spectrometry imaging and develops a new idea for studying drug synergies based on self-immolative structures.

Somatic reversion in Wiskott-Aldrich syndrome: Case reports and mechanistic insights.

This report describes two brothers from India and a Chinese patient with somatic reversion of an inherited deleterious mutation in the WAS gene. Both the Indian siblings had inherited a single nucleotide deletion causing a frameshift mutation (c.1190del, p.Pro397Argfs*48) (variant 1: marked in blue) from the mother. Another variant (variant 2: marked in red), a 12-nucleotide deletion at position 1188-1199 (c.1188_1199del, p.P401_P404del) was also found, which resulted in restoration of the frame and subsequent rescue of the protein sequence. DNA sequencing from buccal mucosal cells revealed only the inherited variant (variant 1), while no reversion mutation was identified in the mucosal cells. Similarly, the Chinese patient was found to have a novel germline 14-base duplication (ACGAAAATGCTTGG) c.120_132 + 1dup (variant 1). This resulted in abolishment of the original splice junction coupled with the creation of a new junction 14 bases 3' and a frameshift mutation with predicted protein truncation p. Thr45Aspfs*. DNA from the patient's PBMC showed co-existence of wild-type and mutated sequences, but only the mutant was present in the buccal cells. Genomic and mRNA analysis of the isolated CD3+ T lymphocytes, CD3- mononuclear cells, and EBV-transformed B lymphocytes indicated that the reverant variant (germline variant was restored to wild-type sequence) were selectively found in CD3+ T lymphocytes.

NK cell membrane/MnO2 hybrid nanoparticle-adjuvanted intranasal vaccines synergistically boost protective immunity against H1N1 influenza infection

Intranasal vaccines hold a huge promise for preventing influenza infection, however, their development is compromised due to mucosal barriers and limited systemic and mucosal immunity. Herein, a novel biomimetic nano-adjuvant based on NK cell membrane (MNK)/MnO2 hybrid nanoparticles (NLipoMn) was developed for intranasal administration of H1N1 inactivated influenza vaccine (IIV). NK cell membrane, served as TLR4 agonist was fused with cationic liposomes encapsulating STING agonist MnO2 NPs (LipoMn) to yield NLipoMn, which inherited versatile characteristics to prolong the nasal retention of IIV. Encouragingly, CCL20 secretion was enhanced by NLipoMn-activated the TLR4/NF-κB pathway in mucosal endothelial cells, thereby promoting submucosal dendritic cells (DCs) recruitment for IIV uptake. Moreover, NLipoMn-IIV synergistically harnessed TLR4 and STING signaling pathway to augment STING activation though NLipoMn-activated the TLR4/NF-κB pathway, which demonstrated a superiority to elicit strong DCs maturation, CD8+ T cells activation, and high levels of antigen-specific IgG, antigen-specific IgA, cytokines secretion, resulting in a robust systemic and mucosal immune responses. Notably, intranasal immunization with NLipoMn-IIV elicited an effective immune protection against homologous and heterologous H1N1 influenza virus infection. This study provides a promising adjuvant candidate to develop needle-free intranasal vaccines that are active against influenza and other respiratory viral infection.

Normalization of CF Immune System Reverses Intestinal Neutrophilic Inflammation and Significantly Improves the Survival of CF Mice

Cystic fibrosis (CF) is an autosomal recessive genetic disorder, affecting multiple organ systems. CF intestinal disease develops early, manifesting as intestinal bacterial overgrowth/dysbiosis, neutrophilic inflammation and obstruction. As unresolvable infection and inflammation reflect host immune deficiency, we sought to determine if the CF-affected immune system plays any significant role in CF intestinal disease pathogenesis. CF and sibling wild-type (WT) mice underwent reciprocal bone marrow transplantation. After immune reconstitution, their mortality, intestinal transit, fecal inflammatory markers, and mucosal immune cell composition were assessed. Moreover, reciprocal neutrophil transfusion was conducted to determine if neutrophil function affects intestinal movement. Furthermore, expression of induced nitric oxide synthase (iNOS) and production of nitric oxide (NO) in CF and WT neutrophils were compared. Lastly, specific iNOS inhibitor 1400W was tested to prevent CF intestinal obstruction. Immune restoration in CF mice reversed the intestinal neutrophilic inflammation, improved the intestinal dysmotility, and rescued the mice from mortality. Transfusion of WT neutrophils into CF mice ameliorated the retarded bowel movement. CF neutrophils expressed significantly more iNOS and produced significantly more NO. Pharmaceutical blocking of iNOS significantly improved intestinal transit and survival of CF mice. CF immune defect plays a critical role in CF intestinal disease development. Activation of iNOS in inflammatory cells produces excessive NO, slows the bowel movement, and facilitates intestinal paralysis and obstruction in CF. Thus, normalization of the CF immune system may offer a novel therapy to treat CF intestinal disease.

Construction of an intelligent pH-sensitive hydrogel drug delivery system and its application in gastric ulcer treatment

The occurrence of gastric ulcer is mainly due to the damage of gastric mucosa which leads to inward flow of gastric acid to corrode the gastric cavity, thus producing gastric fever, vomiting, loss of appetite and other adverse reactions. Prevention and treatment of such injuries are crucial. In this study, a simple and non-toxic composite hydrogel (PLGA/CMCS) was prepared through Schiff base reaction between poly(lactic-co-glycolic acid) (PLGA) and carboxymethyl cellulose (CMCS). Subsequently, compound 1 was loaded onto it to construct PLGA/CMCS@1. A series of structural characterization and performance experiments were conducted on these novel hydrogel polymers. Furthermore, we detected the gene expression level of Fas/FasL apoptotic pathway after treating the cells with different concentrations of PLGA/CMCS@1 through an ethanol-induced damaged gastric mucosal epithelial cell model. The results showed that the system was able to significantly regulate the expression of genes related to the apoptotic pathway. The system can regulate the Fas/FasL apoptosis pathway to treat gastric ulcer.

Sesquiterpene-enriched Extract of Chinese Agarwood (Aquilaria sinensis) Alleviates Bile Reflux Gastritis through Suppression of Gastric Mucosal Cell Apoptosis via the Wnt/β-catenin Signaling Pathway

Ethnopharmacological relevanceChinese agarwood (Aquilaria sinensis) has been a traditional treatment for digestive disorders in South and East Asia. While sesquiterpenes are recognized as the key active constituents of Chinese agarwood, the efficacy and mechanism of the sesquiterpene-enriched extract of Chinese agarwood (PEE) on bile reflux gastritis (BRG) remain unclear. Aim of the studyTo explore the protective impact of PEE against BRG and unveil its underlying mechanism in suppressing apoptosis of gastric mucosal cells. Materials and methodsA taurocholic acid (TCA)-induced BRG mouse model was used to assess PEE's protective effects on gastric mucosa histopathology. Transcriptomic analysis was conducted to identify the key signaling pathways affected by PEE. The impact of PEE on apoptosis modulation and Wnt/β-catenin signaling in GES-1 cells was examined. Additionally, the influence of PEE on the Wnt/β-catenin pathway in BRG mouse gastric mucosa was evaluated. ResultsPEE substantially improved gastric tissue damage and inflammation in BRG mice. Transcriptomic analysis revealed that PEE modulates genes linked to apoptosis and the Wnt/β-catenin pathway. In TCA-induced GES-1 cell, PEE enhanced cell viability and mitigated apoptosis via the Wnt/β-catenin pathway, a process potentially mediated by IWP-2, an antagonist of this pathway. Similar regulatory effects were noted in the gastric mucosa of BRG mice. ConclusionOur research suggests that PEE exerts a protective effect on the gastric tissue through modulating the Wnt/β-catenin pathway to combat apoptosis, which highlights the potential of PEE as a natural remedy for BRG and warrants further investigation into its therapeutic benefits.

CGAS regulates metabolic reprogramming independently of STING pathway in colorectal cancer

BackgroundCyclic GMP-AMP synthase (cGAS) is widely acknowledged for detecting cytosolic chromatin fragments and triggering innate immune responses through the production of the second messenger cGAMP, which subsequently activates the adaptor protein STING. However, the role of cGAS in regulating metabolic reprogramming independently of STING activation has not yet been explored. MethodsGene set enrichment pathway analysis (GSEA) based on TCGA transcriptomics, combined with Seahorse metabolic analysis of CRC cell lines and human normal colonic mucosa cell line FHC, was performed to profile the metabolic features in CRC. cGAS doxycycline- (dox) inducible knockout (iKO) CRC sublines were generated to investigate the role of cGAS in CRC. Transcriptome and proteome data from COAD cohorts were utilized to evaluate the RNA and protein expression levels of cGAS in COAD tissues and normal colon tissues. Overall survival information of patients with COAD was used to evaluate the prognostic value of cGAS expression. Colony formation assays were conducted to evaluate the clonogenicity of CRC cells under different situations. Flow cytometry detecting the signal of fluorogenic reactive oxygen species (ROS) probes was performed to evaluate the total cellular and mitochondrial oxidative stress level in CRC cells. A propidium iodide (PI) staining assay was used to evaluate the cell death level in CRC cells. Quantitative PCR (qPCR) was conducted to detect the RNA level of STING pathway downstream target genes. Mass spectrometry was used for the identification of novel binding partners of cGAS in CRC cells. Co-immunoprecipitation (co-IP) was conducted to confirm the interaction between cGAS and NDUFA4L2. ResultsBy integrating metabolic pathway analysis based on TCGA transcriptomics with Seahorse metabolic analysis of a panel CRC cell lines and the human normal colonic mucosa cell line FHC, we demonstrated that CRC cells exhibit typical characteristics of metabolic reprogramming, characterized by a shift from oxidative phosphorylation (OXPHOS) to glycolysis. We found that cGAS is critical for CRC cells to maintain this metabolic switch. Specifically, the suppression of cGAS through siRNA-mediated knockdown or doxycycline-inducible knockout reversed this metabolic switch, resulting in increased OXPHOS activity, elevated production of OXPHOS byproduct reactive oxygen species (ROS), and consequently caused oxidative stress. This disruption induced oxidative stress, ultimately resulting in cell death and reduced cell viability. Moreover, significant upregulation of cGAS in CRC tissues and cell lines and its association with poor prognosis in CRC patients was observed. Subsequently, we demonstrated that the role of cGAS in regulating metabolic reprogramming does not rely on the canonical cGAS-STING pathway. Co-immunoprecipitation combined with mass spectrometry identified NDUFA4L2 as a novel interactor of cGAS. Subsequent functional experiments, including mitochondrial respiration and oxidative stress assays, demonstrated that cGAS plays a crucial role in sustaining elevated levels of NDUFA4L2 protein expression. The increased expression of NDUFA4L2 is essential for cGAS-mediated regulation of metabolic reprogramming and cell survival in CRC cells. ConclusioncGAS regulates metabolic reprogramming and promotes cell survival in CRC cells through its interaction with NDUFA4L2, independently of the canonical cGAS-STING pathway.

Progress in Transdifferentiation of Autologous Alternative Cell Sources into Corneal Epithelial Cells.

Corneal limbal epithelial stem cells (LESCs) play a crucial role in corneal epithelium regeneration. Severe damage to these cells can result in limbal stem cell deficiency (LSCD), characterized by repeated corneal conjunctivalization, leading to corneal turbidity and scar formation. Restoring functional LESCs and their ecological location are essential for treating LSCD. The goal of this review is to provide researchers and clinicians with key insights into LESCs biology and to conclude the current cell-based therapies advancement in LSCD treatments. Therapeutic cell resources mainly include mesenchymal stem cells (MSCs), embryonic stem cells (ESCs), induced pluripotent stem cells (iPSCs), skin keratinocyte stem cells (SKCs), and oral mucosal epithelial cells (OMECs).

Methanolic Extract of the Nutritional Plant (Diospyros kaki Thunb.) Exhibits Anticancer Activity by Inducing Mitochondrial Dysfunction in Colorectal Cancer Cells.

Background/Objectives:Diospyros kaki, the most widely cultivated species of persimmon, has been long used in traditional medicine since its leaves' extracts contain high amounts of flavonoids and terpenoids, endowed with several beneficial effects. However, its anticancer activity towards colorectal cancer (CRC) has not been investigated in depth. Methods: The effect of a methanolic extract of D. kaki leaves, rich in kaempferol and quercetin derivatives, have been evaluated on an E705 CRC cell line, representative of most CRC patients, and on SW480 cells, carrying a KRAS-activating mutation. Results: This extract is effective in reducing tumor cells' viability without affecting the healthy mucosa cell line CCD 841. In fact, Western blot experiments showed its ability to induce apoptosis in cancer cells by increasing oxidative stress and disrupting mitochondrial functionality, as shown by reactive oxygen species measurement and Seahorse analysis. Conclusions: With the aim of increasing healthspan, as well as the substantial societal and macroeconomic costs associated with cancer, our results could pave the way to a role for D. kaki extract in both CRC treatment and prevention.

Single-cell transcriptomics of rectal organoids from individuals with perianal fistulizing Crohn’s disease reveals patient-specific signatures

Perianal fistulizing Crohn’s disease (CD) is a severe gastrointestinal disorder causing extensive mucosal damage with limited treatment options. Severe manifestations of the disease appear at higher rates in non-Europeans but the genetic and cellular mechanisms driving the disease phenotypes remain poorly understood. Herein, we tested whether pathologic determinants in the epithelial stem cell compartment could be detected at the transcript level in rectal organoids derived from a diverse patient population. Rectal organoid and mucosal cells from endoscopic biopsies of each patient having perianal fistulizing CD or no disease controls were prepared for and sequenced at the single cell level. After cell type annotations based on expressed marker genes, samples were analyzed by principal components, for differential transcript expression, cell type proportions, and pathway enrichment. After QC, we produced 77,044 rectal organoid cells (n = 13 patients; 8 CD, 5 controls) with high quality sequences that identified 10 distinct epithelial subtypes, that we compared to 141,367 mucosal epithelial cells (n = 29 patients; 18 CD, 11 controls). Consistent with mucosal epithelial cells, rectal organoids prominently displayed disease signatures represented by the stem and transit amplifying regions of the rectal crypt, including alterations in transcriptional signatures of metabolic, epigenetic, and proliferating pathways. Organoids also retained their gender- and ancestral-specific gene expression signatures. However, they lacked many of the inflammatory signatures observed in epithelial cells from diseased mucosa. Perianal CD patient derived rectal organoids reflect gene expression signatures related to disease, gender, and ancestry, suggesting they harbor inherent properties amenable to further patient-specific, disease-related experimentation.

Single-cell analysis of nasal epithelial cell development in domestic pigs

The nasal mucosa forms a critical barrier against the invasion of respiratory pathogens. Composed of a heterogeneous assortment of cell types, the nasal mucosa relies on the unique characteristics and complex intercellular dynamics of these cells to maintain their structural integrity and functional efficacy. In this study, single-cell RNA sequencing (scRNA-seq) of porcine nasal mucosa was performed, and nineteen distinct nasal cell types, including nine epithelial cell types, five stromal cell types, and five immune cell types, were identified. The distribution patterns of three representative types of epithelial cells (basal cells, goblet cells, and ciliated cells) were subsequently detected by immunofluorescence. We conducted a comparative analysis of these data with published human single-cell data, revealing consistent differentiation trajectories among porcine and human nasal epithelial cells. Specifically, basal cells serve as the initial stage in the differentiation process of nasal epithelial cells, which then epithelial cells. This research not only enhances our understanding of the composition and transcriptional signature of porcine nasal mucosal cells but also offers a theoretical foundation for developing alternative models for human respiratory diseases.

Nicotine-induced CHRNA5 activation modulates CES1 expression, impacting head and neck squamous cell carcinoma recurrence and metastasis via MEK/ERK pathway

The mucosal epithelium of the head and neck region (including the oral cavity, nasal cavity, pharynx, nasopharynx, and larynx) is the primary site exposed to tobacco smoke, and its presence of nicotinic acetylcholine receptors (nAChRs) has been observed in the mucosal epithelial cells of this area. It remains unclear whether HNSC cells can migrate and invade through nAChR signaling. A model of HNSC cells exposed to nicotine is established. Cell proliferation following nicotine exposure is assessed using the CCK-8 assay, while migration and invasion are evaluated through wound healing and Transwell assays. The effects of CHRNA5 knockdown and overexpression are also investigated. Immunofluorescence staining is used to analyze CHRNA5 expression and localization, and clonogenic assays are performed to measure colony proliferation after CHRNA5 knockdown and overexpression. The interaction between CHRNA5 and CES1 is examined using molecular docking, co-immunoprecipitation, and immunofluorescence. Differentially expressed genes are subjected to pathway enrichment analysis, and MEK/ERK protein expression and phosphorylation are validated via western blot. Tumor formation assays are performed in nude mice using sh-CHRNA5 Cal27 cells, followed by western blot and immunohistochemical staining. Additionally, laryngeal and hypopharyngeal cancer tissues are analyzed through immunohistochemistry. Nicotine significantly enhanced the proliferation, migration, and invasion capabilities of head and neck tumor cells, including Cal27, Fadu, HN6, and Tu686 cells, through the expression of CHRNA5. Knockdown of CHRNA5 can reduce cell migration, invasion, and proliferation, whereas nicotine exposure can reverse this trend. Additionally, the mRNA and protein expression of CES1 decreases with the knockdown of CHRNA5, indicating a regulatory relationship between the two. Transcriptomics revealed that the knockdown of CHRNA5 is associated with the MEK/ERK signaling pathway. Further cellular- and tissue-level evidence confirmed that the levels of p-MEK/MEK, p-ERK/ERK, and CES1 decreased following knockdown of CHRNA5, a trend that nicotine can reverse. Nicotine promotes the proliferation, migration, and invasion of HNSC by upregulating CHRNA5 expression. Knockdown of CHRNA5 reduces these effects, which can be reversed by nicotine. Nicotine exposure activates CHRNA5, regulating CES1 expression via the MEK/ERK pathway, contributing to the recurrence and metastasis of head and neck squamous carcinoma.

Intracellular metabolome elucidates the time-of-day-dependent response to hydrogen peroxide in salmonid gill epithelial cells

The internal timekeeping system regulates the daily cycle of physiological and behavioural changes in living organisms. This rhythmic phenomenon also influences cellular responses to reactive oxygen species, such as hydrogen peroxide (H2O2). However, the temporal interaction between H2O2 and fish mucosal cells is not well understood. This study examined the temporal variations of immunological and physiological responses to H2O2 in salmonid gill cells using the RTgill-W1 cell line. The results showed that gene expression levels varied during a 24-h cycle but did not exhibit rhythmicity. The presence of a 12-h light-dark cycle (12L:12D) signal increased gene expression levels compared to a 24-h dark cycle (0L:24D). To investigate whether the time of day affects the defences in gills, cells were exposed to H2O2 at two different times (Zeitgebertime 2, ZT2, or ZT14). Although significant expression changes were observed in genes related to stress and NF-κB signalling, only a limited time-dependent pattern of response to H2O2 was observed. The intracellular metabolome of gill cells was primarily composed of organic acid and derivatives, organoheterocyclic compounds, benzoids, organic oxygen and nitrogen compounds. Exposure to H2O2 at ZT2 led to significant changes in the metabolome compared to the control group, while no such changes were observed at ZT14. Within the control groups, the concentrations of 11 metabolites significantly varied between ZT2 and ZT14, with higher levels at ZT14. These metabolites were involved in arginine biosynthesis, amino acid metabolism, and nitrogen metabolism. In contrast, the level of 26 metabolites significantly varied between ZT2 and ZT14 in H2O2-exposed groups, with lower levels at ZT14. Comparing control and H2O2-exposed groups at ZT2, 38 metabolites were affected, primarily organic acid and derivatives and organic oxygen compounds. Functional annotation revealed that these altered metabolites were involved in 15 different pathways, with valine, leucine, and isoleucine biosynthesis being the most affected. This study reveals the presence of a time-dependent response to H2O2 in salmonid gill cells, which is reflected in the intracellular metabolome. The findings provide new insights into the temporal regulation of mucosal defences in fish.